The nicotinic acetylcholine receptor (AChR) is a multisubunit membrane protein that mediates synaptic transmission at the neuromuscular junction and is the target of attack in the disease myasthenia gravis. This project will investigate the mechanisms by which this protein is assembled and transported to the cell surface of myotubes. It will also investigate the role of the AChR in the assembly of specialized pre- and postsynaptic structures. A major tool in these investigations will be genetic variants of a mouse muscle cell line that express reduced amount of the AChR on the myotube surface. Two of the variants that we have isolated are deficient in synthesis of the alpha subunit of the AChR. The cause of this defect will be investigated and the use of these variants for in vitro mutagenesis studies on the alpha subunit will be explored. The variants will also be used in co-cultures to determine the properties of nerve-muscle contacts formed in the absence of the AChR. A third variant that makes the AChR, but accumulates it in an intracellular compartment, appears to have a glycosylation defect, and will be used to explore the role of sugars in the transport of the AChR to the cell surface. Finally, an in vitro translation system will be used to investigate the topographical orientation of AChR subunits in membrane and their assembly into the oligomeric AChR.